Amino acid sequence around the thiol and reactive acyl groups of human complement component C4

Area of Science:

• Biochemistry
• Immunology
• Proteomics

Background:

• Complement component 4 (C4) activation by C1s generates reactive acyl and thiol groups.
• These reactive sites are located in the C4d fragment, derived from the alpha'-chain of C4b.
• The native molecule's haemolytic activity is linked to these reactive groups.

Purpose of the Study:

• To elucidate the structural basis of C4 reactivity.
• To determine the location of reactive acyl and thiol groups within the C4d fragment.
• To investigate sequence homology between C4d and other related proteins.

Main Methods:

• Limited digestion of C4b using C3b inactivator and C4-binding protein to isolate C4d.
• Chemical modification of C4d with [1,4-14C]putrescine and iodo[2-14C]acetic acid.
• Peptide generation via CNBr digestion and subsequent sequence analysis.

Main Results:

• An 88-residue sequence from the N-terminus of C4d was established.
• A specific octapeptide containing the reactive thiol and acyl groups was identified.
• This octapeptide exhibited significant sequence homology with equivalent regions in alpha 2-macroglobulin and C3.

Conclusions:

• The identified octapeptide is crucial for the unique reactivity of the thiol ester bond in C4.
• Sequence homology suggests a common structural motif for thiol ester reactivity across C4, alpha 2-macroglobulin, and C3.
• These findings contribute to understanding the molecular mechanisms of complement activation and protein interactions.